1. Field of the Invention
The detection and identification of microorganisms using Raman spectroscopy.
2. Description of the Prior Art
There are many effective methods for the detection of microorganisms. At present, rapid, sensitive tests include fluorescent (fluorescence immunoassay or FIA), or radioactive labels (radio immunoassay or RIA) on the antibody attached in the antigen-antibody complex. Enzymes can be attached to the antibody to produce products which are more easily detected (ELISA). However, such processes (ELISA, RIA and FIA) tend to be labor intensive and not easily adapted to automation. The fluorescence method suffers from background interference and the RIA method is hampered by policies which discourage the use of radioisotopes in routine processes. If only small numbers of bacteria are present separation of the complex from the labelled antibody can be very difficult.
Among the most highly developed of the new rapid detection techniques is mass spectroscopy and its various combinations with gas chromatography (bacterial byproducts from cultures) and pyrolysis methods. Gas chromatography is highly effective in detecting characteristic bacterial metabolic products. Flow cytometry provides for the rapid detection, identification, and separation of cells. Total luminescence spectroscopy can detect organisms very rapidly. The various immunological methods also can be very specific and very rapid. All of these methods have their distinct advantages and disadvantages.
Mass spectroscopy may be unequalled in identification of pure cultures and it is very rapid and sensitive. However, it is expensive to use, requires the destruction of samples, and is of questionable use in the analysis of complex mixtures. Flow cytometry is perhaps even more costly, requires extensive sample preparation, and in many aspects is limited in its scope of applicability. Luminescence techniques are of little use except in studies of pure cultures unless combined with immunological methods. Immunological methods are unequalled in specificity and speed, as well as sensitivity. Yet, they are often impractical to use unless very expensive and perishable materials are available in a state of constant readiness. Such methods are not practical for a wide range of organisms. Gas chromatography requires that cells be grown and, hence, this method is generally slow and of limited applicability.
In bacterial analysis normally the cost effective means of analysis involves isolating organisms and then growing them in controlled cultures. This process is very slow and relatively labor intensive.
It is known to detect and identify microorganisms based on resonance Raman spectra, U.S. Pat. No. 4,847,198. A beam of visible or ultraviolet light energy contacts a microorganism under investigation. A portion of the light energy is absorbed by the microorganism and a portion of the light energy is ‘emitted’ from the microorganism at a lower energy level. The emitted light energy (resonance enhanced Raman scattering) can be correlated to a specific microorganism.